Substituted 4-(anilinomethylene)-3-galanthamaninones

ABSTRACT

This disclosure described compounds of the class of substituted 4-(anilinomethylene)-3-galanthamaninones useful as antibacterial agents and central nervous system depressants.

United States Patent Morlock et al.

June 27, 1972 SUBSTITUTED 4- (ANILlNOMETHYLENE)-3- GALANTHAMANINONESElizabeth Benn Morlock, Bronx; Leon Goldman, Nanuet, both of NY.

Inventors:

Appl. No.: 34,591

US. Cl. ..260/240.3, 260/310 R, 260/3462 M, 424/285 Int. Cl. ..C07d99/04 Field of Search ..260/240.3, 346.2 M

[56] References Cited OTHER PUBLICATIONS Barton et al., J. Chem. Soc.,806- 17(1962).

Primary ExaminerHenry R. Jiles Assistant Examiner-G. Thomas ToddAttorney-Edward A. Conroy, Jr.

[57] ABSTRACT This disclosure described compounds of the class ofsubstituted 4-(anilinomethylene)-3-galanthamaninones useful asantibacterial agents and central nervous system depressants.

8 Claims, No Drawings SUBSTITUTED 4-(ANILINOMETHYLENE)-3-GALANTI'IAMANINONES BRIEF SUMMARY OF THE INVENTION CH2CH2 N-CH; I CH2Wherein R is phenyl, p-trifluoromethylphenyl, trifluoromethylphenyl oro-trifiuoromethylphenyl.

DETAILED DESCRIPTION OF THE INVENTION The novel compounds of the presentinvention are, in general, yellow or orange crystalline solids withcharacteristic melting points and spectral properties. The free basesare soluble in such common organic solvents as lower alkanols,chloroform, methylene chloride, acetone, benzene, N,N-dimethylformamide, dioxane and dimethyl sulfoxide. They are, however,generally insoluble in water.

The organic free .bases of this invention form non-toxic acid-additionsalts with a variety of organic and inorganic saltforming reagents.Thus, acid-addition salts, formed by admixture of the organic free basewith an equivalent amount of an acid, suitably in a neutral solvent, areformed with such acids as sulfuric, phosphoric, hydrochloric,hydrobromic, citric, lactic, tartaric, acetic, gluconic, ascorbic, andthe like. For purposes of this invention, the organic free bases areequivalent to their non-toxic acid-addition salts.

The novel compounds of the present invention may be readily preparedfrom 3-galanthamaninonc (I) in accordance with the following reactionscheme:

0 o H l wherein R is a lower alkyl group having up to four carbon ethylformate ethanol (III) to pH 7.0

atoms, M is an alkali metal, and R is as hereinabove defined. Thestarting material (I) is B-galanthamaninone (galanthaminone ornarwedine) which has been described by D. H. R. Barton and G. W. Kirby,J. Chem. Soc., 806 (1962) and by G. Combes and J. C. Lefebvre, Bull.Soc. Chim. France, 1805 (1962). Condensation of 3-galanthamaninone (I)with ethyl formate in benzene in the presence of an alkali metalalkoxide (ROM) at room temperature for a period of time of 2-3 daysaffords the corresponding 4-(hydroxymethylene)-3-galanthamaninone alkalimetal enolate 'salt (II). Suitable alkali metal alkoxides which may beemployed in this condensation are, for example, sodium methoxide,potassium ethoxide, lithium tert-butoxide, and the like. Dissolution ofthe water soluble 4-(hydroxymethylene)-3- galanthamaninone alkali metalenolate salts (II) in water followed by acidification to pH 7.0 affordsthe 4-(hydroxymethylene)-3-galanthamaninone (III) as the water solubleinner salt. Both 4-(hydroxymethylene)-3-galanthamaninone (III) and itsalkali metal enolate salts (II) are new compounds and are includedwithin the purview of the present invention.

Treatment of 4-(hydroxymethylene)-3-galanthamaninone (III) with anaromatic amine of the formula R-NH (wherein R is as hereinabove defined)in ethanol at the reflux temperature for a few hours affords thecorresponding 4- (anilinomethylene)-3-galanthamaninone (IV). Similarly,treatment of a 4-( hydroxymethylene)-3-galanthamaninone alkali metalenolate salt (II) with an aromatic amine of the formula RNH in glacialacetic acid at room temperature for 10-20 hours also affords thecorresponding 4- (anilinomethylene )-3-galanthamaninone (IV).

Certain of the novel compounds of the present invention (when R is o-,m-, or p-trifluoromethylphenyl) are useful as antibacterial agents andpossess broad-spectrum antibacterial activity in vitro against a varietyof standard laboratory microorganisms as determined by the agar-dilutionstreakplate technique. In this assay, the compounds to be tested aremade up to contain 2.5 mg. of test compound per milliliter of solution.Observing sterile techniques, two-fold serial dilutions are made of eachtest solution. One milliliter of each of the original solutions and ofeach of the serial dilutions is then added to 9 ml. of warm sterilenutrient agar capable of supporting growth of the bacterial testcultures. The standard sterile nutrient agar solutions containing thedifferent dilutions of the test compounds, along with suitable andcomparable control dilutions containing no test compound, are thenallowed to cool in Petri dishes thereby forming solidified agar plates.The test bacteria are prepared for use by growing in broth overnight. Aloopful of each of the live suspensions in sterile physiological salinesolution is then, still employing sterile techniques, streaked upon thesurfaces of each of the agar plates and the resulting streaked platesare then incubated. After an appropriate period of time, each of thestreaks on each of the plates is inspected visually and the extent, ifany, of bacterial growth is noted. The minimal inhibitory concentration(expressed in micrograms per milliliter) is defined as the concentrationof test compound causing complete inhibition of growth of any particularorganism. In a representative operation, and merely by way ofillustration, the minimal inhibitory concentration of 4-[(a,a,a-trifluoro-ptoluidino)methylene]-3 galanthamaninone (l) and4-[(a,a,atrifluoro-m-toluidino)methylene]-3-galanthamaninone (2) againsta variety of test organisms as determined in the abovedescribed assayare set forth in the following table:

Minimal inhibitory conc. (meg/ml.)

Organism Clostridium sporogene: ATCC 7955 Certain ofthe novel compoundsof the present invention are valuable central nervous system depressantsof low toxicity and were shown to possess CNS depressant activity asdetermined by animal experiments as follows. The compound wasadministered intraperitoneally in a 2 percent starch vehicle to groupsof six mice at three or more graded dose levels. At 15- minute and30-minute intervals after treatment, each animal was placed on themidpoint of a horizontal steel rod 1.55cm. in diameter and about 6 drn.in length), positioned 45.7 cm. above the surface of the table, andforced to walk toward a platform at either end of the rod. The criterionof inability to perform this act was consistent slipping to the side orfalling off the rod. The effective dose for reduced rod-walking ability(RWD was calculated or approximated from the data, and the time of peakeffect was estimated from the data. One-half of the RWD dose was givenintraperitoneally to each mouse in groups of five. At the time of peakeffect, as determined above, each group of mice was put into theactophotometer for a period of 5 minutes and the motor activity countswere recorded and compared to controls. The compound was administered toadditional groups of five mice at graded doses and tested similarly. Thedose (MDD that caused a 50 percent reduction in motor(Anilinomethylene)-3-galanthamaninone and pyrazolo-[5,4':3,4]galanthamanine were shown to induce ataxia (RWD at a dose of 250and 270 mg./kg. of bodyweight, respectively, and to reduce locomotoractivity MDD at a dose of 122 and 1 l6 mgJkg. of body weight,respectively.

When mixed with suitable excipients or diluents, the compounds of thisinvention can be prepared as pills, capsules, tablets, powders,solutions, suspensions, and the like for unit dosage and to simplifyadministration.

The invention will be described in greater detail in conjunction withthe following specific examples.

EXAMPLE 1 600 ml. of dry benzene is added 2079 g. of sodium methoxideactivity was estimated. 4-

and the mixture is allowed to stir for 0.5 hour. Ethyl formate (30 ml.)is added and the deep orange mixture is stirred at room temperature for3 days. The mixture is filtered and the filtrate is evaporated todryness under reduced pressure to yield4-(hydroxymethylene)-3galanthamaninone sodium salt as a thick red syrup.The syrup is dissolved in 200 ml. of icewater and the pH of the solutionis adjusted to 7.0 by the addition of glacial acetic acid. The resultingsolution is extracted successively with methylene chloride andchloroform. During the extraction process a tan solid precipitates fromthe aqueous solution and this is collected by filtration to yield 8.05g. of

4-(hydroxymethylene)-3-galanthamaninone as a tan solid,

m.p. 144-149 C. The methylene chloride and chloroform extracts areevaporated to dryness under reduced pressure to yield a thick red syrupwhich solidifies on trituration with anhydrous ether. Filtration gives11.8 g. of crude 4-(hydroxymethylene)-3-galantha.maninone.

EXAMPLE 2 Preparation of 4-(anilinomethylene)-3-galanthamaninone Asuspension of 0.100 g. of 4-(hydroxymethylene)-3- galanthamaninone and0.03 ml. (0.033 g.) of aniline in 10 ml. of absolute ethanol is refluxedfor 2 hours. The mixture is cooled to room temperature and filtered toyield 0.054 g. of 4- (anilino methylene)-3-galahthamaninone as yellowneedles, m.p. 2l6-218 C. (dec.); lt 229 nm (22,000), 257 nm (15,700) and377 nm (21,000).

EXAMPLE 3 Preparation of 4-(anilinomethylene )-3-galanthamaninone To asuspension of 2.50 g. of 4-(hydroxymethylene )-3- galanthamaninonesodium salt in 75 ml. of glacial acetic acid is added 0.73 ml. (0.74g.)of aniline and the mixture is stirred at room temperature for 16 hours.The solution is evaporated under reduced pressure to an orangesemi-solid residue. A solution of the residue in 75 ml. of water is madealkaline by addition of concentrated ammonium hydroxide and theresulting yellow precipitate is collected by filtration. Crystallizationfrom absolute ethanol yields 1.68 g. of 4-(anilinomethylene)-3-galanthamaninone as yellow crystals, m.p. 2l62l8 C. (dec.).

EXAMPLE 4 Preparation of 4-[(a,u,a-trifluoro-p-toluidino)methylene]-3-galanthamaninone A suspension of 1.00 g. of 4-(hydroxymethylene)-3-galanthamaninone and 0.518 g. of p-aminobenzotrifluoride in 25 ml. ofabsolute ethanol is refluxed for 2 hours. The solvent is removed underreduced pressure to yield a deep red syrup as residue. On triturationwith ether an insoluable tan solid results which is removed byfiltration and the filtrate is evaporated to dryness under reducedpressure to yield a deep red syrup as residue. This residue ischromatographed on 15 g. of alumina (activity Ill). The column is elutedwith benzene and twelve lO-ml. cuts are collected, and then withbenzeneether (3:1 and nine 10-ml. cuts are collected. Cuts 10-22 arecombined and evaporated to dryness under reduced pressure to yield 0.230g. of crude 4-[(a,a,a-trifluoro-ptoluidino)methylene]-B-gaIanthamaninoneas yellow crystals, m.p. l82l 85 C. Recrystallization from absoluteethanol gives 0.140 g. of 4-[(a,a,a-trifluoro-p-toluidino)methylene]'3-galanthamaninone as yellow crystals, m.p. l85l8.' C.; A 228 nm(620,800), 262 nm (16,200) and 370 nm (22,800).

EXAMPLE 5 Preparation of 4-[(a,a,a-trifluoro-o-toluidino)methylene]-3-galanthamaninone The procedure of Example 4 is repeated, substituting anequimolar amount of o-aminobenzotrifluoride for thepaminobenzotrifluoride employed in that example. There is thus obtainedthe 4-[(a,a,a-trifluoro-o-toluidino)methylene1 3-glaanthamaninone.

EXAMPLE 6 max.

EXAMPLE 7 Preparation of pyrazolo[ 5 ,4 3,4] galanthamanine A solutionof 2.00 g. of 4-(hydroxymethylene)-3- galanthamaninone sodium salt and0.32 ml. of hydrazine hydrate in 50 ml. of glacial acetic acid isallowed to stir at room temperature for 18 hours. The solvent is removedby evaporation under reduced pressure to yield an orange semisolidresidue. A solution of the residue in water is chilled and made alkalinewith concentrated ammonium hydroxide and the resultant orangeprecipitate is collected by filtration, washed well with water andair-dried. The crudesolid is recrystallized from 30 ml. of 50 percentaqueous ethanol to yield 104g. of pyrazolo[5,4':3,4]galanthamanine aspale yellow needles, m.p. 140-150 090132230 nm 10,200 258 nm (5,100) and280 nm' (1,700). Pyrazolo[5',4:-3,4] galanthamanine may be representedby the following tautomeric formulas:

The free base forms non-toxic acid-addition slats with a variety oforganic and inorganic salt-forming reagents. Thus, acid-addition salts,formed by admixture of the free base with one or two equivalents of anacid, suitably in a neutral solvent, are formed with such acids assulfuric, phosphoric, hydrochloric, hydrobromic, citric, lactic,tartaric, acetic, gluconic, ascorbic, and the like.

We claim:

l. A compound selected from the group consisting of those of theformula:

CHaO

wherein R is selected from the group consisting of[phenyl,]otrifluoromethylphenyl, m-trifluoromethylphenyl andptrifluoromethylphenyl; and the non-toxic pharmaceutically acceptableacid-addition salts thereof.

2. A compound according to claim 1 wherein R is otrifluoromethylphenyl.

3. A compound according to claim 1 wherein R is mtrifluoromethylphenyl.

A compound according to claim 1 wherein R is ptrifluoromethylphenyl.

5. The compound 4-(hydroxymethylene )-3- galanthamaninone represented bythe formula:

6. A compound of the formula:

\CH2CH2

2. A compound according to claim 1 wherein R is o-trifluoromethylphenyl.
 3. A compound according to claim 1 wherein R is m-trifluoromethylphenyl.
 4. A compound according to claim 1 wherein R is p-trifluoromethylphenyl.
 5. The compound 4-(hydroxymethylene)-3-galanthamaninone represented by the formula:
 6. A compound of the formula:
 7. A compound according to claim 6 wherein M is sodium; 4-(hydroxymethylene)-3-galanthamaninone sodium enolate salt.
 8. A compound according to claim 6 wherein M is potassium; 4-(hydroxymethylene)-3-galanthamaninone potassium enolate salt. 